Pertussis (whooping cough) is a vaccine-preventable but re-emerging, highly infectious respiratory disease caused by Bordetella pertussis. There are currently no effective treatments for pertussis, complicating care for non-vaccinated individuals, especially newborns. Disease manifestations are predominantly caused by pertussis toxin (PT), a pivotal virulence factor classified as an ADP-ribosylating AB-type protein toxin. In this work, an unbiased approach using peptide libraries, bioassay-guided fractionation and mass spectrometry revealed α₁-antitrypsin (α₁AT) as a potent PT inhibitor. Biochemistry-, cell culture- and molecular modeling-based in vitro experimentation demonstrated that the α₁AT mode of action is based on blocking PT-binding to the host target cell surface. In the infant mouse model of severe pertussis, α₁AT expression was reduced upon infection. Further, systemic administration of α₁AT significantly reduced B. pertussis-induced leukocytosis, which is a hallmark of infant infection and major risk factor for fatal pertussis. Taken together our data demonstrates that α₁AT is a novel PT inhibitor and that further evaluation and development of α₁AT as a therapeutic agent for pertussis is warranted. Importantly, purified α₁AT is already in use clinically as an intravenous augmentation therapy for those with genetic α₁AT deficiency and could be repurposed to clinical management of pertussis.